Cystic fibrosis (CF) is a common, lethal genetic disease caused by dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel. The commonest mutation, F508del-CFTR disrupts CFTR processing and intracellular transport. It also impacts on channel stability and gating at the cell surface, with the result that F508del-CFTR Cl- channels are slow to open, but rundown rapidly at 37 oC (1, 2). Here we investigate the effects of temperature on F508del-CFTR channel gating to understand better its instability. To address our aim, we studied CFTR Cl- channels in excised inside-out membrane patches from C127 mouse mammary epithelial cells stably expressing wild-type and F508del- human CFTR at 23, 27, 30, 33 and 37 oC. The pipette (external) solution contained 10 mM Cl-, whereas the bath (internal) solution contained 147 mM Cl-, ATP (1 mM) and protein kinase A (PKA; 75 nM) at 23 – 37 oC; voltage was -50 mV. At 23 oC, the single-channel current amplitudes of wild-type and F508del-CFTR were -0.54 ± 0.03 and -0.50 ± 0.01 pA (means ± SEM; n ≥ 4; p > 0.05; student’s unpaired t-test). As temperature was elevated to 37 oC, the current amplitude of wild-type and F508del-CFTR increased linearly to -0.75 ± 0.04 and -0.74 ± 0.02 pA (n ≥ 4; p > 0.05; student’s unpaired t-test). However, temperature had different effects on the open probability (Po) of wild-type and F508del-CFTR. For wild-type CFTR, Po increased exponentially from 0.24 ± 0.02 at 23 oC to 0.46 ± 0.04 at 37 oC (n ≥ 4; student’s unpaired t-test). By contrast, for F508del-CFTR, Po increased from 0.06 ± 0.01 at 23 oC to 0.09 ± 0.01 at 30 oC before falling back to 0.06 ± 0.01 at 37 oC (n ≥ 4). To understand better the temperature-dependence of Po, we performed an analysis of bursts. For wild-type CFTR over the temperature range 23 – 37 oC, mean burst duration (MBD) decreased 2-fold and interburst interval (IBI) fell 7-fold (n = 4). However, for F508del-CFTR, MBD decreased 4-fold and IBI fell 7-fold over the temperature range 23 – 37 oC (n = 4). In conclusion, our data suggest that temperature has similar effects on the single-channel current amplitudes of wild-type and F508del-CFTR. However, the gating behaviour of the two channels responds differently to temperature. For wild-type CFTR, maximal activity occurs at 37 oC, whereas for F508del-CFTR, it is around 30 oC. Thus, these data suggest that temperature might have different effects on the conduction and gating properties of CFTR.